The role of NF-kappaB during the PMA-induced megakaryocytic differentiation of K562 cells was investigated using K562 cells transfected with each or both subunits of NF-kappaB. The NF-kappaB subunit-transfected cells have shown much higher sensitivity to PMA-induced differentiation than their parental cells. This result was consistent with the findings that PMA-stimulated activities of NF-kappaB were markedly increased in the NF-kappaB subunit-transfected cells in comparison with their parental cells and PMA-induced differentiation was enhanced by pretreatment with IkappaB-alpha antisense oligonucleotide in the NF-kappaB subunit-transfected cells. Meanwhile, there were basically no difference in the basal and PMA-stimulated MAP kinase activities among the parental and NF-kappaB subunit-transfected cells, respectively. However, PMA-induced differentiation was blocked by pretreatment with PD98059, a specific inhibitor of MEK, in both parental and NF-kappaB-transfected cells. Therefore, these results suggest that during the PMA-induced megakaryocytic differentiation of K562 cells, NF-kappaB works downstream of MAP kinase, or that activation of both NF-kappaB and MAP kinase pathways is involved.